The rank of reference genes for 'stomach cancer cell lines' was identical with that from the geNorm analysis, but was slightly different in other categories.
As a result, use of these two pairs of co-regulated genes did not affect the final ranking of the reference genes by using geNorm software.
The ranking of the reference genes was almost identical to that determined by the geNorm software.
To overcome this circular problem, Vandesompele et al. [ 33] first developed more than ten years ago a method called geNorm, which allows the evaluation and ranking of candidate reference genes in terms of expression stability (or suitability as normalizing gene).
This algorithm calculates the average pair wise variation of a given candidate reference genes set with all other genes under evaluation and assigns a measure of its expression stability (M), based on which a ranking of candidate reference genes is produced [ 8].
In caprine samples, the ranked lists of reference genes for samples and analyses differed.
For PBMC samples, the ranked lists of reference genes, produced by geNorm and NormFinder were the same and the most stable reference gene was ACTB.
Also, the ranked list of reference genes, provided by both software, was the same for PBMC and for whole blood samples.
Then, alignment results are converted into a format that is accepted by variationBlast, which reports a ranked list of reference regions that show homology to the read and annotate the SVs for each region as follows.
For those genomes with good correlation coefficients between the relative rank order of reference species and number of contributed interactions (> 0.9), these data are virtually linear (for an example see Figure 3c), with 115 such genomes having slopes greater than 1.0 (average slope 4.75, see Additional File 3).
